Antimicrobial amphiphiles and methods for their use

ABSTRACT

The present invention is directed to compositions useful for antimicrobial applications. These compositions comprise amphiphilic compounds.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is the National Stage of International Application No.PCT/US2011/057036, filed Oct. 20, 2011, which claims the benefit of U.S.Provisional Application No. 61/394,938, filed Oct. 20, 2010, thedisclosures of which are incorporated herein by reference in theirentireties.

TECHNICAL FIELD

The present invention is directed to compositions useful forantimicrobial applications. These compositions comprise amphiphiliccompounds.

BACKGROUND

Antibiotic resistance, particularly in human pathogens such asmethicillin-resistant Stapyhlococcus aureus (MRSA), Clostridiumdifficile, vancomycin-resistant enterococci, and Mycobacteriumtuberculosis, has increased dramatically in the last 20 years, which hasthreatened the ability to treat hospital- and community-acquiredinfections. Recently, widespread resistance to antibacterial compoundssuch as triclosan has also been observed, necessitating newantimicrobial compounds, particularly novel antibiotics and antiseptics.In addition to developing new antimicrobials by modifying existingdrugs, antimicrobials with novel structures must also be developed,particularly compounds that will be difficult for organisms toinactivate or to resist via mutation. Compounds that present a uniquestrategy of activity are of premium importance.

Thus, new compounds having anti-microbial properties are needed.

SUMMARY

The present invention is directed to pharmaceutical compositionscomprising at least one compound of formula I:

wherein R₁, R₂, R₃, R₅, R₆, R₇, R₈, R₉, and R₁₀ are each independentlyC₁₋₁₈alkyl or R₁, R₂, and R₃, and/or R₅, R₆, and R₇, and/or R₈, R₉, andR₁₀, together with the nitrogen atom to which they are attached, form apyridinium or substituted pyridinium; m is 1; n is 0 or 1; p is 0 or 1;t is 0 or 1; R₄ is C₈₋₂₂alkyl; and X is halogen or tartrate; and apharmaceutically acceptable carrier or diluent. Methods of inhibitingbacterial growth using pharmaceutical compositions and compounds of theinvention are also described.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1. Depicts antimicrobial activity of preferred embodiments of theinvention against. (A) S. aureus, (B) E. faecalis, and (C) E. coli.Filled symbols (♦, ▪, ●) indicate the MIC and MBC for each organism,while open symbols of the same shape (⋄, □, ∘) indicate the timenecessary to kill 100% of the organism. *=time necessary to kill was >72h.

FIG. 2. Depicts antimicrobial activity of preferred embodiments of theinvention against. (A) S. aureus, (B) E. faecalis, and (C) E. coli.Filled symbols (♦, ▪, ●) indicate the MIC and MBC for each organism,while open symbols of the same shape (⋄, □, ∘) indicate the timenecessary to kill 100% of the organism. *=time necessary to kill was >72h. †=MIC values>500 μM.

DETAILED DESCRIPTION OF ILLUSTRATIVE EMBODIMENTS

Compounds demonstrating anti-bacterial properties have been developed.These compounds are useful alone or in pharmaceutical compositions forthe inhibition of bacterial growth.

The pharmaceutical compositions of the invention comprise at least onecompound of formula I:

wherein

-   -   R₁, R₂, R₃, R₅, R₆, R₇, R₈, R₉, and R₁₀ are each independently        C₁₋₁₈alkyl or R₁, R₂, and R₃, and/or R₅, R₆, and R₇, and/or R₈,        R₉, and R₁₀, together with the nitrogen atom to which they are        attached, form a pyridinium or substituted pyridinium;    -   m is 1;    -   n is 0 or 1;    -   p is 0 or 1;    -   t is 0 or 1;    -   R₄ is C₈₋₂₂alkyl; and    -   X is halogen or tartrate;    -   and a pharmaceutically acceptable carrier or diluent.

Certain embodiments of the invention include pharmaceutical compositionswherein m is 1; n is 0; p is 0; and t is 1. In such embodiments, it ispreferred that each of R₁, R₂, and R₃ is independently C₁₋₃alkyl. Morepreferably, each of R₁, R₂, and R₃ is methyl. In such embodiments,preferred compounds of formula I have the following structure:

Other preferred embodiments of the invention include pharmaceuticalcompositions wherein m is 1; n is 1; p is 0; and t is 1. In suchembodiments, the compound of formula I preferably has one of thefollowing structures:

In such embodiments wherein m is 1; n is 1; p is 0; and t is 1, it ispreferred that each of R₁, R₂, R₃, R₅, R₆, and R₇ are independentlyC₁₋₃alkyl. More preferred are embodiments wherein each of R₁, R₂, R₃,R₅, R₆, and R₇ is methyl.

In other embodiments, R₁, R₂, and R₃ and R₅, R₆, and R₇, together withthe nitrogen atom to which they are attached, form a pyridinium. Ofthese embodiments, preferred compounds of formula I for use inpharmaceutical compositions of the invention include:

Other preferred embodiments include those wherein R₁, R₂, and R₃ and R₅,R₆, and R₇, together with the nitrogen atom to which they are attached,form a pyridyl-substituted pyridinium.

In other embodiments of the invention, R₁, R₂, and R₃ are eachindependently C₁₋₃alkyl and R₅, R₆, and R₇, together with the nitrogenatom to which they are attached, form a pyridinium or substitutedpyridinium.

All of the foregoing embodiments of the invention include an R₄ moiety.For all such embodiments, R₄ is preferably —C₁₀H₂₁. Also preferred areembodiments where R₄ is —C₁₂H₂₅. Other preferred embodiments includethose wherein R₄ is —C₁₄H₂₉. Particularly preferred are embodimentswherein R₄ is —C₁₆H₃₃. Additional embodiments include those wherein R₄is —C₁₈H₃₇.

Preferred compounds of the invention for use in pharmaceuticalcompositions of the invention include:

Other preferred compounds of formula I for use in pharmaceuticalcompositions of the invention include those selected from the followingTable 1:

TABLE 1

Comp. No. Substitution R₄ 1 2, 3 —C₁₄H₂₉ 2 2, 4 —C₁₄H₂₉ 3 2, 5 —C₁₀H₂₁ 42, 5 —C₁₂H₂₅ 5 2, 5 —C₁₄H₂₉ 6 2, 5 —C₁₆H₃₃ 7 2, 5 —C₁₈H₃₇ 8 2, 6 —C₁₄H₂₉9 3, 5 —C₁₄H₂₉ 10  3, 5 —C₁₆H₃₃

Additionally preferred compounds of formula I for use in pharmaceuticalcompositions of the invention include those selected from the followingTable 2:

TABLE 2

Comp. No. Substitution R₄ 15 2, 3 —C₁₄H₂₉ 40 3, 5 —C₁₄H₂₉ 16 3, 5—C₁₆H₃₃

A preferred compound of formula I for use in pharmaceutical compositionsof the invention is:

Also within the scope of the invention are pharmaceutical compositionsof the invention that include compounds of formula I wherein m is 1; nis 1; p is 0; and t is 0. In such embodiments, it is preferred that thecompound of formula I is one of

In such embodiments, it is preferred that R₁, R₂, R₅ and R₆ are eachC₁₋₃alkyl; and R₃ and R₇ are each independently C₈₋₁₈alkyl.

In other such embodiments, it is preferred that R₁, R₂, R₅ and R₆ areeach methyl; and R₃ and R₇ are each independently C₁₂₋₁₄alkyl.

In still other such embodiments, it is preferred that R₁ and R₂ are eachC₁₋₃alkyl; R₃ is C₈₋₁₈alkyl; and R₅, R₆, and R₇, together with thenitrogen atom to which they are attached, form pyridinium.

In yet other such embodiments, it is preferred that R₁ and R₂ are eachmethyl; and R₃ is C₁₂₋₁₄alkyl.

Preferred compounds of such embodiments include:

Other embodiments of the invention include pharmaceutical compositionsthat include compounds of formula I wherein m is 1; n is 1; p is 1; andt is 0. In such embodiments, it is preferred that R₁, R₂, R₅, R₆, R₈ andR₉ are each independently C₁₋₃alkyl; and R₃, R₇, and R₁₀ are eachindependently C₈₋₁₈alkyl.

In other such embodiments, it is preferred that R₁, R₂, R₅, R₆, R₈ andR₉ are each methyl; and R₃, R₇, and R₁₀ are each independentlyC₁₂₋₁₄alkyl.

Preferred compounds for use in pharmaceutical compositions of theinvention include

In yet other such embodiments, it is preferred that R₁, R₂, R₅, and R₆,are each independently C₁₋₃alkyl; R₃ and R₇ are each independentlyC₈₋₁₈alkyl; and R₈, R₉, and R₁₀, together with the nitrogen atom towhich they are attached, form pyridinium.

Preferred compounds for use in pharmaceutical compositions of theinvention include those in the following table:

Comp. No. R₃ R₇ 25 —C₈H₁₇ —C₈H₁₇ 26 —C₁₀H₂₁ —C₁₀H₂₁ 27 —C₁₂H₂₅ —C₁₂H₂₅28 —C₁₄H₂₉ —C₁₄H₂₉ 29 —C₁₆H₃₃ —C₁₆H₃₃

In all of the foregoing embodiments of the invention, it is preferredthat X is Br.

Also within the scope of the invention are methods of inhibitingbacterial growth comprising contacting a bacteria with any of thecompounds of the invention described herein. Preferred bacteria includeStaphylococcus aureus, Entercoccus faecalis, Escherichia coli,Pseudomonas aeruginosa or a combination thereof.

As used herein, “alkyl” refers to a straight or branched chainhydrocarbon having from one to 22 carbon atoms. Examples of alkyl groupsinclude methyl, ethyl, propyl, isopropyl, butyl, isobutyl, tert-butyl,pentyl, isopentyl, hexyl, and the like.

As used herein, “pyridinium” refers to the cationic form of pyridine.

As used herein, “substituted pyridinium” refers to the cationic form ofpyridine wherein one or more carbon atoms of the pyridine ring issubstituted with, for example, C₁₋₆alkyl, C₆₋₁₀aryl, such as phenyl ornaphthyl, or a heteroaryl such as pyridyl.

As used herein, “halogen” refers to F, Cl, Br, and I.

As used herein, “tartrate” refers to the negatively charged form oftartaric acid.

The applicable carrier or diluent may be selected on the basis of thechosen route of administration and standard pharmaceutical practice asdescribed, for example, in Remington's Pharmaceutical Sciences (MackPub. Co., Easton, Pa., 1985), the disclosure of which is herebyincorporated by reference in its entirety. Suitable examples of liquidcarriers and diluents include water (particularly containing additivesas above, e.g. cellulose derivatives, preferably sodium carboxymethylcellulose solution), alcohols (including monohydric alcohols andpolyhydric alcohols, e.g. glycols) and their derivatives, and oils (e.g.fractionated coconut oil and arachis oil), or mixtures thereof. Thecompounds of the invention may be administered in an effective amount byany of the conventional techniques well-established in the medicalfield. The compounds employed in the methods of the present inventionincluding, for example, the compounds of formula I may be administeredby any means that results in the contact of the active agents with theagents' site or sites of action in the body of a patient. The compoundsmay be administered by any conventional means available.

Compounds of the invention can be prepared according to methods known inthe art, for example, as described in K. Z. Roszak et al. Biscationicbicephalic (double-headed)amphiphiles with an aromatic spacer and asingle hydrophobic tail, J. Colloid and Interface Science 331 (2009)560-564, the entirety of which is incorporated herein. Exemplary methodsfor preparing certain embodiments are the invention are describedherein. Those skilled in the art can readily modify the proceduresdescribed herein to prepared compounds within the full scope of theinvention.

Accordingly, a series of commercially available dimethylphenols as wellas p-cresol were exposed to Williamson ether synthesis conditions,resulting in phenol alkylation in high yields (compound A, 72-99%,Scheme 1 and accompanying table). Light-promoted benzylic bromination(NBS, benzoyl peroxide) subsequently generated the correspondingbenzylic bromides B in low yields. Exposure to trimethylamine in ethanolat reflux completed the syntheses of these amphiphiles (C), which werepurified by recrystallization (EtOH/H₂O) in good yield.

Com- Chain % yield % yield of % yield of pound m Substitution length (R)of A bromide B compound 1 2 2,3 C₁₄H₂₉ 78% 39% 68% 2 2 2,4 C₁₄H₂₉ 84%31% 11% 3 2 2,5 C₁₀H₂₁ 90% 14% 76% 4 2 2,5 C₁₂H₂₅ 88% 25% 81% 5 2 2,5C₁₄H₂₉ 86% 23% 87% 6 2 2,5 C₁₆H₃₃ 72% 28% 78% 7 2 2,5 C₁₈H₃₇  78%* 25%83% 8 2 2,6 C₁₄H₂₉ 99% 8% 40% 11 1 para (4) C₁₀H₂₁ 92% 30% 76% 12 1 para(4) C₁₄H₂₉ 80% 15% 74% *= acetone used as solvent.

Compounds 9 and 10 can be prepared via an alternate method, as benzylicbromination of the 3,5-dimethylalkoxyphenol derivatives yielded acomplex mixture of products. Compounds 9 and 10 were prepared from5-hydroxyisophthalate dimethylester (14, Scheme 2). Accordingly,Williamson ether synthesis (tetradecyl or hexadecyl bromide, K₂CO₃,CH₃CN, reflux) followed by lithium aluminum hydride reduction yieldedthe diol (D). Subsequent reaction with phosphorus tribromide providedbis-benzylic bromides B, which were transformed to the correspondingtrimethylammonium cations in good yield as described above.

Other compounds of the invention, for example, compounds 21 to 29 can beprepared according to the sequence set forth in Scheme 3.

Other compounds of the invention, for example, compounds 30, 31, 32, 35,36, and 37 can be prepared according to the sequence set forth in Scheme4.

Other compounds of the invention, for example, compound 41, can beprepared according to the sequence set forth in Scheme 5.

The compounds of the invention are useful for inhibiting the growth ofbacteria by contacting a bacteria with a compound of formula I.Preferably, the compound of formula I is formulated into apharmaceutical composition for use in inhibiting bacterial growth. Theinvention is particularly useful in inhibiting the growth of, forexample, Staphylococcus aureus, Entercoccus faecalis, Escherichia coli,Pseudomonas aeruginosa or a combination thereof.

The growth of other bacteria can also be inhibited using the compoundsand compositions of the invention, for example, Salmonella species,Shigella species, Streptococcus pyogenes, and Haemophilus influenzae.

All compounds except for 2,5-C10 (3) and SDS were effective at killingS. aureus at low micromolar concentrations, with compounds 2,3-C14 (1),2,4-C14 (2), 3,5-C14 (9), MC10 (11), and MC14 (12) effective at <10 μM.against E. faecalis, compounds 1, 2, and 5-12 had MIC and MBC values at16-50 μM, with 4 effective at 125 μM. The concentration of compoundsnecessary to kill E. coli were between those necessary to kill E.faecalis and P. aeruginosa, generally between 31-63 μM.

Compounds 2,4-C14 (2) and 3,5-C14 (9) had the lowest MIC and MBC valuesagainst all 4 strains, with 3,5-C16 (10) and MC10 (11) being nearly aseffective across the strains. The monocationic compounds MC10 (11) andMC14 (12) were much more effective at inhibiting Gram-positive organismsthan Gram-negatives.

In addition to determining MIC and MBC values, compounds were assayedfor the time necessary to kill 2.5×10⁶ cfu/mL. The time kill resultswere similar for S. aureus and E. faecalis. 2,5-C18 (7), 3,5-C16 (10),and MC14 (12) each killed within 15 min, while MC10 (11) killed within30 min. E. coli was killed by 3,5-C16 (10) within 15 min, while 2,3-C14(1), 2,5-C18 (7), and MC10 (11) killed within 30 min Although a numberof compounds were effective at killing within 72 h, clearly 3,5-C16 (10)was the most efficient at killing the 3 strains tested, while 2,5-C18(7) was nearly as efficient. MC14 (12) was very effective against theGram-positive strains, but ineffective against E. coli.

Additionally, aliquots of cells killed by the compounds were placed onslides and visualized microscopically to determine if any intact cellswere present after treatment. Visual inspection suggested that theeffective compounds kill via cell lysis. MICs and MBCs of preferredcompounds of the invention are set forth in Table 3. Time killing ofbacterial strains by preferred compounds of the invention are set forthin Table 4.

TABLE 3 MICs and MBCs (μM) of Compounds of the Invention S. aureus E.faecalis P. aeruginosa Compound (G⁺) (G⁺) E. coli (G⁻) (G⁻) 2,3-C14 (1)6 31 63 125 2,4-C14 (2) 6 16 31 125 2,5-C10 (3) 250 500 >500 >5002,5-C12 (4) 31 125 500 >500 2,5-C14 (5) 16 31 63 250 2,5-C16 (6) 16 5063 125 2,5-C18 (7) 16 31 63 250 2,6-C14 (8) 16 50 125 500 3,5-C14 (9) 831 31 >63 3,5-C16 (10) 16 31 63 >63 MC10 (11) 8 16 63 250 MC14 (12) 616 >500 >500 SDS (13) 250 500 >500 >500 2,3-C14pyr (15) 4 8 16 633,5-C14pyr (40) 4 16 16 63 2,5-C14dipyr (41) 16 31 16 125 3,5-C16pyr(16) 16 31 31 125 mX-14,14 (31) 4 6 47 125 pX-14,14 (32) 16 16 63 125M-P,12,12 (27) 2 2 4 8 M-P,14,14 (28) 8 8 16 125 M-P,16,16 (29) 31 31 63125 M-14,14,14 (23) 31 31 63 125 G⁺ = Gram-positive, G⁻ = Gram-negative

TABLE 4 Time Killing of Bacterial Strains by Compounds of the Invention.Data are reported as time (h) when all cells were killed by 100 μM ofcompound. Compound S. aureus (G⁺) E. faecalis (G⁺) E. coli (G⁻) 2,3-C14(1) 1 24 0.5 2,4-C14 (2) 1 2 1 2,5-C10 (3) NK NK NK 2,5-C12 (4) NK NK NK2,5-C14 (5) 48 48 72 2,5-C16 (6) 24 NK 2 2,5-C18 (7) 0.25 0.25 0.52,6-C14 (8) 48 NK NK 3,5-C14 (9) 1 1 1 3,5-C16 (10) 0.25 0.25 0.25 MC10(11) 0.5 0.5 0.5 MC14 (12) 0.25 0.25 NK NK = 100% of cells not killedafter treatment. Pseudomonas aeruginosa was not tested because the MICvalues for all compounds were greater than 100 μM. SDS was not usedbecause it was not effective at 100 μM.

Also within the scope of the invention are pharmaceutical compositionscomprising a compound of formula IA:

-   -   wherein    -   R₁, R₂, and R₃ are each independently C₁₋₆alkyl or R₁, R₂, and        R₃, together with the nitrogen atom to which they are attached,        form a pyridinium;    -   m is 1 or 2;    -   R₄ is C₈₋₂₂alkyl; and    -   X is halogen or tartrate;    -   and a pharmaceutically acceptable carrier or diluent.

Preferred embodiments include pharmaceutical compositions wherein mis 1. Also within the scope of the invention are pharmaceuticalcompositions, wherein the compound of formula IA has the followingstructure:

Other preferred embodiments include pharmaceutical compositions whereinm is 2.

Preferred pharmaceutical compositions comprise one or more of thefollowing compounds:

-   -   wherein        -   R₁, R₂, and R₃ are each independently C₁₋₆alkyl or R₁, R₂,            and R₃, together with the nitrogen atom to which they are            attached, form a pyridinium;        -   m is 1 or 2;        -   R₄ is C₈₋₂₂alkyl; and        -   X is halogen or tartrate;    -   and a pharmaceutically acceptable carrier or diluent.

Preferred embodiments of the invention include compounds of formula IA,wherein each of R₁, R₂, and R₃ is C₁₋₃alkyl. In other embodiments, eachof R₁, R₂, and R₃ is methyl. In yet other embodiments, R₁, R₂, and R₃,together with the nitrogen atom to which they are attached, form apyridinium.

Other preferred embodiments of the invention include compounds offormula IA, wherein R₄ is C₁₀₋₁₈alkyl. Yet other embodiments includecompounds wherein R₄ is —C₁₀H₂₁. In still other embodiments, R₄ is—C₁₂H₂₅. In still other embodiments are compounds of formula IA, whereinR₄ is —C₁₄H₂₉. Also preferred are compounds of formula IA, wherein R₄ is—C₁₆H₃₃. Preferred pharmaceutical compositions also include compounds offormula IA, wherein R₄ is —C₁₈H₃₇.

Also within the scope of the invention are pharmaceutical compositions,wherein X is Br.

Preferred pharmaceutical compositions of the invention include compoundsof formula IA is selected from the following Table:

m Substitution R₄ 2 2, 3 —C₁₄H₂₉ 2 2, 4 —C₁₄H₂₉ 2 2, 5 —C₁₀H₂₁ 2 2, 5—C₁₂H₂₅ 2 2, 5 —C₁₄H₂₉ 2 2, 5 —C₁₆H₃₃ 2 2, 5 —C₁₈H₃₇ 2 2, 6 —C₁₄H₂₉ 2 3,5 —C₁₄H₂₉ 2 3, 5 —C₁₆H₃₃ 1 4 —C₁₀H₂₁ 1 4 —C₁₄H₂₉

Other preferred pharmaceutical compositions of the invention includethose, wherein the compound of formula IA is selected from the followingTable:

m Substitution R₄ 2 2, 3 —C₁₄H₂₉ 2 3, 5 —C₁₆H₃₃

Within the scope of the invention are methods of inhibiting bacterialgrowth comprising contacting a bacteria with a compound of formula IA.In preferred embodiments, the bacteria is Staphylococcus aureus,Entercoccus faecalis, Escherichia coli, Pseudomonas aeruginosa or acombination thereof.

Also within the scope of the invention are compounds of formula IB:

-   -   wherein        -   R₁, R₂, and R₃ are each independently C₁₋₆alkyl or R₁, R₂,            and R₃, together with the nitrogen atom to which they are            attached, form a pyridinium;        -   m is 1 or 2; wherein when m is 1, the —CH₂—[N(R₁)(R₂)(R₃)]            group is at the 2, 3, 5, or 6 position of the phenyl ring            and when m is 2, the —CH₂—[(N(R₁)(R₂)(R₃)] groups are not at            the 2,5 positions of the phenyl ring;        -   R₄ is C₈₋₂₂alkyl; and        -   X is halogen or tartrate.

The following experimental details are exemplary only and are notintended to limit the scope of the invention.

Experimental Section

Bacterial Strains and Culture Conditions.

Bacterial strains Staphylococcus aureus subsp. aureus ATCC® 29213™,Entercoccus faecalis ATCC® 29212™, Escherichia coli ATCC® 25922™, andPseudomonas aeruginosa strain Boston 41501 ATCC® 27853™ were obtainedfrom the American type Culture Collection (ATCC, Manassas, Va., USA). S.aureus and E. faecalis are Gram-positive pathogens, P. aeruginosa is aGram-negative pathogen, and E. coli is a non-pathogenic Gram-negativeorganism. All strains are reference strains for the Clinical andLaboratory Standards Institute (CLSI) for antimicrobial susceptibilitytesting of nonfastidious bacteria, and were grown in Mueller-Hintonbroth at 37° C. as recommended (Wayne, Pa. National Committee forClinical Laboratory Standards (2009) Methods for Dilution AntimicrobialTests for Bacteria That Grow Aerobically—Approved Standard M07-A8).

Broth Microdilution MIC and MBC Determination.

The broth microdilution for determining the MIC and MBC of antimicrobialcompounds was performed as previously described in Wayne, supra.Briefly, all compounds were serially diluted and 100 μl of each dilutionwas added to microtiter plate wells in triplicate. Overnight cultures ofbacterial cells were diluted to a final inoculum of approximately 5×10⁶cfu/mL, and 100 μl of this suspension was added to all wells yielded5×10⁵ cfu/well. Cell concentrations and viability were verified byserial dilution and plating for each experiment. Microtiter plates wereincubated at 37° C. for 72 h. The MIC was determined as the lowestconcentration of compound to completely inhibit growth as detected bythe unaided eye. From each set of triplicate wells, 100 μl was thenplated on Todd-Hewitt agar (THA, Becton, Dickinson and Company, Sparks,Md., USA) and incubated for 24 h and examined to determine the MBCvalues for each compound. The MBC was determined as the lowestconcentration of compound at which there were no colonies growing on theplate. MIC and MBC experiments were performed a minimum of 3 times foreach organism.

Bacterial Killing Assays.

To determine the relative temporal effectiveness of the synthesizedamphiphiles, overnight cultures were diluted in broth to a concentrationof 2.5×10⁶ cfu/ml. Compounds were added to the cultures to a finalconcentration of 100 μM, and tubes were incubated at room temperature.At 15, 30, 60, 90, 120, 180 min, 24, 48, and 72 h, one-hundredmicroliter aliquots were plated on THA plates, and the plates wereincubated overnight at 37° C. Data reported are times when no colonieswere observed growing on plates after overnight incubation.Additionally, 100 μl of cells treated with compounds for 72 h wereplaced on slides, Gram-stained, and visualized microscopically todetermine if intact cells were still present after treatment.

MIC values for the compounds of the invention were determined by thebroth microdilution method as described herein. MICs were determined asthe lowest concentration which prevented visible growth at 72 h. Aftermicrotiter plates were incubated and MICs determined, aliquots from eachof the wells were placed on TSA plates and incubated to determine theMBC values for each of the compounds. The MBC was determined as thelowest concentration of compound at which there were no colonies growingon the plate. The MBC and MIC values were identical in every case,indicating that the compounds are effective at killing the bacteria, notjust inhibiting growth. SDS was used for comparison in these studies.For the MICs and MBCs, there was variability both with various compoundsagainst individual strains, and with a single compound across strains.

Synthesis and Analysis.

All solvents and reagents were used as received from the indicatedchemical supplier unless otherwise specified. Melting points for solidswere measured using a Mel-Temp apparatus with a digital thermometer(uncorr). Nuclear magnetic resonance spectra were recorded using one ofthe following instruments, as noted: 600 (¹H: 600 MHz, ¹³C: 150 MHz) or400 (¹H: 400 MHz, ¹³C: 100 MHz). The solvent residual peak was used as areference. Coupling constants are estimated to be correct within ±0.1Hz. High-resolution mass spectra (HRMS) were recorded on a AccuTOFtime-of-flight mass spectrometer using either a Direct Analysis in RealTime (DART) interface or electrospray ionization (ESI) interface, asnoted, in positive ion mode.

Synthesis of 1-8, 11, 12.

Compounds 1-8, 11, and 12 were prepared via a three step synthesisstarting from the corresponding methyl phenol derivative as shown inScheme S1 using General protocols A-C, described below. Specificsynthetic details for each derivative not previously reported arepresented subsequently.

Synthesis of 9, 10.

Compounds 9 and 10 were prepared via a four step synthesis starting fromdimethyl-5-hydroxyisophthalate (14) as shown in Scheme S1 using Generalprotocols D, E, F and C, described below. Specific synthetic details foreach derivative are presented subsequently.

General Protocol A: Williamson Ether Synthesis Starting fromX,Y-Dimethyl Phenol or p-Cresol [Y.-Z. Lee, X., Chen, S.-A. Chen, P.-K.Wei, W.-S. Fann, J. Am. Chem. Soc. 2001, 123, 2296].

The alkyl bromide, phenol derivative and potassium carbonate werecombined in a round bottom flask in acetonitrile or acetone, as noted.The flask was equipped with a stir-bar, and a water-cooled condenser andprotected under a nitrogen atmosphere. The mixture was heated at refluxfor one to three days. Synthetic progress was monitored by thedisappearance of the signal (triplet) from the C1 hydrogens on the alkylbromide (˜3.5 ppm) in ¹H NMR. Upon completion, the reaction was removedfrom heat and excess K₂CO₃ was removed by gravity filtration (rinsedwith CH₂Cl₂). The crude product was concentrated in vacuo and theresulting material was dissolved in CH₂Cl₂, washed with 1 M NaOH (2×, toremove unreacted phenol derivative), dH₂O (2×) and brine (1×), driedover Na₂SO₄, gravity filtered, and concentrated in vacuo to remove thesolvent. If needed, the product was purified by column chromatography(as noted). The material was of sufficient purity (by ¹H and ¹³C NMR) tobe used in the subsequent reactions.

General Protocol B: Benzylic Bromination [Y.-Z. Lee, X., Chen, S.-A.Chen, P.-K. Wei, W.-S. Fann, J. Am. Chem. Soc. 2001, 123, 2296].

All glassware was dried in an oven overnight and flushed with N₂ (g)prior to use. A solution of the synthesized aryl alkyl ether (A1-A8, A11or A12, as noted) in carbon tetrachloride was prepared in a round bottomflask equipped with a water-cooled condenser, magnetic stir bar andprotected with under a nitrogen atmosphere. N-bromosuccinimide (NBS) andbenzoyl peroxide (cat) were added and the reaction was either heated toreflux on an oil bath or reacted at room temperature under irradiationfrom a 300 W/82V tungsten halogen lamp (as noted). Reaction progress wasmonitored by the disappearance of the insoluble NBS at the bottom of theflask and the appearance of the lower density insoluble succinimidebyproduct that floated to the surface of the reaction mixture. Afterfiltration of the succinimide, the solvent was removed in vacuo. Thecrude product was recrystallized from n-hexane.

General Protocol C: Menchutkin Reaction.

The bis-benzylbromide derivate (B1-B10, as noted) or benzyl bromidederivative (B11 or B12, as noted) was suspended in absolute ethanol in atwo-neck round-bottom flask which was attached to a nitrogen (g)protected water-cooled condenser. Trimethylamine (33 wt. % in ethanol)was added via syringe, the second neck was sealed with a glass stopperand the reaction was heated to reflux on an oil bath. The solid startingmaterial dissolved upon heating. After at least four hours under reflux,excess NMe₃ and EtOH were allowed to evaporate under a flow of N₂ gas byremoving the stopper from the second neck. The residual material wastaken up in absolute EtOH, transferred to a clean round bottom flask andthe solvent was removed in vacuo. The product was recrystallized fromEtOH/Et₂O. Residual volatile material was removed by vacuum drying overP₂O₅ at 80-115° C.

General Protocol D: Williamson Ether Synthesis Starting fromdimethyl-5-hydroxyisophthalate.

The alkyl bromide, dimethyl-5-hydroxyisophthalate (14) and potassiumcarbonate were combined in a round bottom flask in acetonitrile. Theflask was equipped with a stir-bar, and a water-cooled condenser andprotected under a nitrogen atmosphere. The mixture was heated at refluxfor one to three days. Synthetic progress was monitored by thedisappearance of the signal (triplet) from the Cl hydrogens on the alkylbromide (˜3.5 ppm) in ¹H NMR. Upon completion, the reaction was removedfrom heat and excess K₂CO₃ was removed by gravity filtration (rinsedwith CH₂Cl₂). The crude product was concentrated in vacuo and theresulting material was dissolved in CH₂Cl₂, washed with 1 M NaOH (2×, toremove unreacted 14), dH₂O (2×) and brine (1×), dried over Na₂SO₄,gravity filtered, and concentrated in vacuo to remove the solvent. Ifneeded, the product was purified by column chromatography (as noted).The material was of sufficient purity (by ¹H and ¹³C NMR) to be used inthe subsequent reactions.

General Protocol E: Ester Reduction.

All glassware was dried in an oven overnight and flushed with N₂ (g)prior to use. A solution of diester D9 or D10 (1 equiv.) in dry Et₂O wasprepared in a 100-mL round bottom flask. In a second round bottom flaskequipped with a magnetic stir bar and a water-cooled condenser, lithiumaluminum hydride (2.2 equiv, 95% Acros) was suspended in dry Et₂O. Theethereal diester solution was added dropwise to the reaction flask viasyringe. The reaction was run under reflux for 3 hours and subsequentlyquenched with the addition of 5 mL NaOH (1M), and 20 mL H₂O. The Et₂Owas removed from the reaction flask via rotary evaporation. The productwas purified by recrystallization from EtOH and trituration from benzeneand hexanes.

General Protocol F: Substitution Using Phosphorus Tribromide.

Phosphorus tribromide (PBr₃) (2.2 equiv, 99% Acros) was added viasyringe to a THF solution of bis-benzyl alcohol E10 or E11 (1 equiv) ina round bottom flask equipped with nitrogen and a magnetic stir bar.After three days at room temperature, the reaction mixture wasconcentrated via rotary evaportaion. The resulting brown liquid waspurified by column chromatography on silica using hexanes followed by anincreasing gradient to EtOAc/hexanes (98/2). The desired product wasobtained after removal of the solvent in vacuo.

1-tetradecyloxy-2,3-dimethylbenzene (A1)

The product was generated via general protocol A using1-bromotetradecane (45.0 mL, 147.9 mmol), 2,3-dimethylphenol (20.15 g,163.3 mmol), K₂CO₃ (81.76 g, 591.6 mmol) and CH₃CN (200 mL). Yielded36.6 g (77.7%) of a brown liquid. ¹H NMR (CDCl₃, 400 MHz) δ: 7.03 (t,³J=7.8 Hz, 1H, Ar—H), 6.76 (d, ³J=7.4 Hz, 1H, Ar—H), 6.70 (d, ³J=8.1 Hz,1H, Ar—H), 3.93 (t, ³J=6.5, 2H, O—CH ₂), 2.27 (s, 3H, Ar—CH ₃), 2.16 (s,3H, Ar—CH ₃), 1.79 (p, ³J=6.9, 2H, O—CH₂—CH ₂), 1.48 (p, ³J=7.1, 2H,O—CH₂—CH₂—CH ₂), 1.27 (m, 22H), 0.88 (t, ³J=6.3, 3H, CH₂—CH ₃). ¹³C NMR(CDCl₃, 100 MHz) δ: 157.20, 137.86, 125.84, 125.33, 122.08, 108.98,68.23, 32.07, 29.86, 29.83, 29.78, 29.61, 29.58, 29.54, 26.33, 22.88,20.17, 14.24, 11.70.

1-tetradecyloxy-2,3-bis(bromomethyl)benzene (B1)

The product was generated via general protocol B using1-tetradecyloxy-2,3-dimethylbenzene (A1, 25.1 g, 78.1 mmol),N-bromosuccinimide (99%, 28.2 g, 158 mmol), benzoyl peroxide (38 mg,0.16 mmol) and CCl₄ (850 mL). The reaction was run in 10 equal batchesat room temperature under irradiation for ˜60 min per batch. After tworecrystallizations of the combined crude product from n-hexane 14.87 g(39.46%) of pure product was obtained as an off-white solid,m.p.=55.7-56.5° C. ¹H NMR (CDCl₃, 300 MHz) δ: 7.24 (t, ³J=8.0 Hz, 1H,Ar—H), 6.94 (d, ³J=7.5 Hz, 1H, Ar—H) 6.84 (d, ³J=8.2, 1H, Ar—H), 4.78(s, 2H, Ar—CH ₂), 4.62 (s, 2H, Ar—CH ₂), 4.02 (t, ³J=6.4, 2H, O—CH ₂),1.84 (p, 2H, O—CH₂—CH ₂), 1.54 (p, 2H, O—CH₂—CH₂—CH ₂), 1.26 (m, 22H),0.88 (t, ³J=6.68, 3H, CH₂—CH ₃). ¹³C NMR (CDCl₃, 150 MHz) δ: 157.65,138.25, 129.94, 125.53, 122.76, 112.33, 109.56, 68.64, 32.28, 30.14,29.74, 29.64, 29.39, 26.11, 23.88, 22.70, 14.21.

[(2-tetradecyloxy-o-phenylene)dimethylene]bis[trimethylamonium bromide](1) (2,3-C14)

The product was generated via general protocol C using1-tetradecyloxy-2,3-bis(bromomethyl)benzene (B1, 4.84 g, 10.2 mmol),NMe₃ (33% wt solution in EtOH, 9.70 mL, 40.6 mmol) and EtOH (100 mL).After recrystallization (EtOH/Et₂O) and drying the reaction produced4.09 g (67.7%) of a white solid. ¹H NMR (CD₃OD, 300 MHz) δ: 7.71 (t,³J=8.1 Hz, 1H, Ar—H), 7.43 (d, ³J=8.4 Hz, 1H, Ar—H), 7.37 (d, ³J=8.1 Hz,1H, Ar—H) 4.93 (s, 2H, Ar—CH ₂), 4.85 (s, 2H, Ar—CH ₂), 4.16 (t, ³J=6.8Hz, 2H, O—CH₂), 3.17 (s, 18H, N—CH ₃), 1.9 (p, ³J=6.9 Hz, 2H, O—CH₂—CH₂), 1.28 (m, 23H), 0.90 (t, ³J=6.5, 3H, CH₂—CH ₃). ¹³C NMR (CD₃OD, 150MHz) δ: 159.70, 132.70, 130.70, 127.20, 117.69, 115.53, 69.52, 64.85,59.18, 52.31, 52.08, 31.71, 29.45, 29.16, 29.05, 28.74, 25.80, 22.36,13.13. Anal. Calcd for C₂₈H₅₄Br₂N₂O.H₂O: C, 54.90; H, 9.21; N, 4.57.Found: C, 54.89; H, 9.31; N, 4.48.

1-tetradecyloxy-2,4-dimethylbenzene (A2)

The product was generated via general protocol A using1-bromotetradecane (44 mL, 0.149 mol), 2,4-dimethylphenol (19.5 mL,0.164 mol), K₂CO₃ (82.4 g, 0.596 mol) and CH₃CN (200 mL). Yielded 43.13g (90.8%). ¹H NMR (CDCl₃, 600 MHz) δ: 7.00 (s, 1H, Ar—H), 6.94 (d,³J=8.41 Hz, 1H, Ar—H), 6.75 (d, ³J=8.15 Hz, 1H, Ar—H), 3.97 (t, ³J=6.46Hz, 2H, OCH ₂), 2.31 (s, 3H, Ar—CH ₃), 2.26 (s, 3H, Ar—CH ₃), 1.83 (p,³J=7.3 Hz, 2H, O—CH₂—CH ₂), 1.52 (p, ³J=7.6 Hz, 2H, O—CH₂—CH₂—CH ₂),1.44-1.28 (m, 22H), 0.95 (t, ³J=7.05, 3H, CH₂—CH ₃). ¹³C NMR (150 MHz)δ: 155.24, 131.45, 129.20, 126.88, 126.66, 111.08, 68.17, 53.41, 32.02,31.90, 29.97, 29.79, 29.78, 29.76, 29.52, 29.50, 29.46, 26.24, 22.78,20.47, 16.19, 14.19.

1-tetradecyloxy-2,4-bis(bromomethyl)benzene (B2)

The product was generated via general protocol B using1-tetradecyloxy-2,4-dimethylbenzene (A2, 20.2 g, 63.4 mmol),N-bromosuccinimide (99%, 22.8 g, 127 mmol), benzoyl peroxide (0.314 g,1.30 mmol) and CCl₄ (680 mL). The reaction was run in 8 equal batches atroom temperature under irradiation for 30-60 min per batch. After tworecrystallizations of the combined crude product from n-hexane 6.66 g(22.3%) of pure product was obtained as a blue solid, m.p.=62.9° C. ¹HNMR (CDCl₃, 400 MHz) δ: 7.36 (d, ³J=2.3 Hz, 1H, Ar—H), 7.30 (dd, ³J=2.3,⁴J=8.5, 1H, Ar—H), 6.82 (d, ³J=8.5, 1H, Ar—H), 4.53 (s, 2H, Ar—CH ₂—Br),4.47 (s, 2H, Ar—CH₂—Br), 4.03 (t, ³J=6.3 Hz, 2H, OCH ₂), 1.83 (p, ³J=7.0Hz, 2H, OCH₂CH ₂), 1.49 (p, ³J=7.5 Hz, 2H, OCH₂CH₂CH ₂), 1.36-1.26 (m,22H), 0.88 (t, ³J=7.0 Hz, 3H, CH₂CH ₃).

[(6-tetradecyloxy-m-phenylene)dimethylene]bis[trimethylamonium bromide](2) (2,4-C14)

The product was generated via general protocol C using1-tetradecyloxy-2,4-bis(bromomethyl)benzene (B2, 4.00 g, 8.40 mmol),NMe₃ (33% wt solution in EtOH, 12.0 mL, 50.4 mmol) and EtOH (200 mL).After recrystallization (EtOH/Et₂O) and drying the reaction produced4.99 g of a white solid. Anal. Calcd for C₂₈H₅₄Br₂N₂O.2H₂O: C, 53.33; H,9.27; N, 4.44. Found: C, 53.35; H, 9.16; N, 4.34.

A3-A7, B3-B7, 3-7 (2,5-C10-2,5-C18).

Synthetic details and analyses of these compounds have previously beenpublished. [Roszak, K. Z.; Torcivia, S. L.; Hamill, K. M.; Hill, A. R.;Radloff, K. R.; Crizer, D. M.; Middleton, A. M.; Caran, K. L. J. ColloidInterface Sci., 2009, 331, 560-5641.]

1-tetradecyloxy-2,6-dimethylbenzene (A8)

The product was generated via general protocol A using1-bromotetradecane (44.7 mL, 0.147 mol), 2,6-dimethylphenol (20.0 g,0.162 mol), K₂CO₃ (81.3 g, 0.588 mol) and CH₃CN (150 mL). Yielded 46.5 g(99.3%) of a yellow/orange liquid. ¹H NMR (CDCl₃, 400 MHz) δ: 7.01 (d,⁴J=7.3 Hz, 2H, Ar—H), 6.91 (t, ³J=6.8 Hz, 3H, Ar—H), 3.76 (t, ³J=6.7 Hz,2H, OCH ₂), 2.28 (s, 6H, Ar—CH ₃), 1.81 (p, ³J=7.1 Hz, 2H, OCH₂CH ₂),1.50 (p, ³J=7.4 Hz, 2H, OCH₂CH₂CH ₂), 1.36-1.27 (m, 22H), 0.89 (t,³J=6.9 Hz, 3H, CH₂CH ₃). ¹³C NMR (CDCl₃, 100 MHz) δ: 156.08, 130.95,128.71, 123.54, 77.32, 77.00, 76.68, 72.28, 31.93, 30.44, 29.70, 29.68,29.66, 29.63, 29.57, 29.37, 26.16, 22.69, 16.26, 14.12.

1-tetradecyloxy-2,6-bis(bromomethyl)benzene (B8)

The product was generated via general protocol B using1-tetradecyloxy-2,6-dimethylbenzene (A8, 32.5 g, 102 mmol),N-bromosuccinimide (99%, 36.3 g, 204 mmol), benzoyl peroxide (0.494 g,2.04 mmol) and CCl₄ (1105 mL). The reaction was run in 13 equal batchesunder irradiation for 30-60 min per batch. After four recrystallizationsof the combined crude product from n-hexane, 8.314 g (18.5%), of thepure product was obtained as a white solid, mp=59.3-59.8° C. ¹H NMR(CDCl₃, 400 MHz) δ: 7.37 (d, ³J=7.62 Hz, 2H, Ar—H), 7.11 (t, ³J=7.65,1H, Ar—H), 4.55 (s, 4H, Ar—CH ₂—Br), 4.10 (t, ³J=6.62 Hz, 2H, OCH ₂),1.90 (p, ³J=7.54 Hz, 2H, OCH₂CH ₂), 1.55 (p, ³J=7.35 Hz, 2H, OCH₂CH₂CH₂), 1.43-1.22 (m, 24H), 0.88 (t, ³J=6.97 Hz, 3H, CH₂CH ₃). ¹³C NMR (150MHz) δ: 155.73, 132.26, 132.10, 132.03, 129.06, 124.92, 124.19, 73.37,31.96, 30.35, 29.74, 29.72, 29.70, 29.65, 29.63, 29.53, 29.41, 28.68,27.76, 26.06, 25.96, 22.73, 16.36, 14.18.

[(2-tetradecyloxy-m-phenylene)dimethylene]bis[trimethylamonium bromide](8) (2,6-C14)

The product was generated via general protocol C using1-tetradecyloxy-2,6-bis(bromomethyl)benzene (B8, 8.00 g, 16.7 mmol),NMe₃ (33% wt solution in EtOH, 24.0 mL, 100 mmol) and EtOH (400 mL).After recrystallization (EtOH/Et₂O) and drying the reaction produced5.53 g (55.6%) of a white solid, mp=221.9-224.9° C. (dec). ¹H NMR (MeOD,600 MHz) δ: 7.81 (d, ³J=7.70 Hz, 2H, Ar—H), 7.48 (t, ³J=7.70 Hz, 1H,Ar—H), 4.58 (s, 4H, Ar—CH ₂—N), 3.93 (t, ³J=7.13 Hz, 2H, O—CH ₂), 3.18(s, 18H, N—CH ₃), 1.98 (p, ³J=7.62 Hz, 2H, O—CH₂—CH ₂), 1.53-1.23 (m,24H), 0.89 (t, ³J=7.0 Hz, 3H, CH₂—CH ₃). ¹³C NMR (150 MHz) δ: 158.12,136.68, 124.05, 121.64, 75.72, 62.45, 50.85, 30.16, 28.52, 27.90, 27.88,27.86, 27.82, 27.57, 24.24, 20.82, 11.53. Anal. Calcd forC₂₈H₅₄Br₂N₂O.H₂O: C, 54.90; H, 9.21; N, 4.57. Found: C, 55.35; H, 9.18;N, 4.58.

Dimethyl-5-tetradecyloxyisophthalate (D9)

The product was generated using general protocol D usingdimethyl-5-hydroxyisophthalate (1.037 g, 4.93 mmol), 1-bromotetradecane(1.34 mL, 4.48 mmol), K₂CO₃ (2.48 g, XXmol), CH₃CN (50 mL) yielded 1.21g (60.2%) of a white solid. ¹H NMR (CDCl₃, 400 MHz) δ: 8.2607 (s, 1H,Ar—H), 7.7396 (s, 2H, Ar—H), 4.0315 (t, 2H, O—CH ₂, ³J=6.62 Hz), 3.9374(s, 6H, COOCH ₃), 1.8046 (p, 2H, O—CH₂—CH ₂ ³J=6.82 Hz), 1.4660 (p, 2H,O—CH₂—CH₂—CH ₂, ³J=6.86 Hz), 1.2582 (m, 22H), 0.8809 (t, 3H, CH₂—CH ₃,³J=6.69 Hz

1-Tetradecyloxy-3,5-bis(hydroxymethyl)benzene (E9)

The product was generated using general protocol E usingdimethyl-5-tetradecyloxyisophthalate (D9, 1.171 g, 2.88 mmol), lithiumaluminum hydride (0.253 g, 6.34 mmol), Et₂O (40 mL) yielded 0.538 g(53.3%) of a white solid. ¹H NMR (CDCl₃, 300 MHz) δ: 6.9336 (s, 1H,Ar—H), 6.8501 (s, 2H, Ar—H), 4.6821 (s, 4H, Ar—CH ₂), 3.9708 (t, 2H,O—CH ₂, ³J=6.58 Hz), 1.7784 (p, 2H, O—CH₂—CH ₂, ³J=7.16 Hz), 1.6328 (t,2H, O—H), 1.4483 (p, 2H, O—CH₂—CH₂—CH ₂, ³J=7.58 Hz), 1.2625 (m, 20H),0.8810 (t, 3H, CH₂—CH ₃, ³J=6.83 Hz).

1-tetradecyloxy-3,5-bis(bromomethyl)benzene (B9)

The product was generated via general protocol F using1-tetradecyloxy-3,5-bis(hydroxymethyl)benzene (E9, 2.79 g, 7.95 mmol),PBr₃ (1.51 mL, 15.9 mmol), and THF (80 mL). After column chromatography(hexanes, then ramped to 2% EtOAc in hexanes), the reaction yielded 1.26g (46%) of a white solid. ¹H NMR (CDCl₃, 600 MHz) δ: 6.9819 (s, 1H,Ar—H), 6.8531 (s, 2H, Ar—H), 4.4279 (s, 4H, Ar—CH ₂), 3.9540 (t, 2H,O—CH₂, ³J=6.55 Hz), 1.7738 (p, 2H, O—CH₂—CH ₂, ³J=7.26 Hz), 1.4474 (p,2H, O—CH₂—CH₂—CH ₂, ³J=7.70 Hz), 1.2631 (m, 22H), 0.8809 (t, 3H, CH₂—CH₃, ³J=7.06 Hz).

[(5-tetradecyloxy-m-phenylene)dimethylene]bis[trimethylamonium bromide](9) (3,5-C14)

The product was generated via general protocol C using1-tetradecyloxy-3,5-bis(bromomethyl)benzene (B9, 0.888 g, 1.86 mmol),NMe₃ (33% wt solution in EtOH, 1.78 mL, 7.46 mmol) and EtOH (20 mL).After recrystallization (EtOH/Et₂O) and drying the reaction produced0.973 g (88.4%) of a white solid, mp=197.2-197.7° C. (dec). ¹H NMR(MeOD, 400 MHz) δ: 7.4055 (s, 1H, Ar—H), 7.3314 (s, 2H, Ar—H), 4.6073(s, 4H, Ar—CH ₂), 4.1045 (t, 2H, O—CH ₂, ³J=6.45 Hz), 3.1796 (s, 18H,N—CH ₃) 1.8241 (p, 2H, O—CH₂—CH ₂, ³J=7.46 Hz), 1.5040 (p, 2H,O—CH₂—CH₂—CH ₂, ³J=7.39 Hz), 1.2936 (m, 20H), 0.9020 (t, 3H, CH₂—CH ₃,³J=7.04 Hz). Anal. Calcd for C₂₈H₅₄Br₂N₂O.H₂O: C, 54.90; H, 9.21; N,4.57. Found: C, 54.96; H, 9.24; N, 4.58.

Dimethyl-5-hexadecyloxyisophthalate (D10)

The product was generated using general protocol D usingdimethyl-5-hydroxyisophthalate (3.920 g, 18.7 mmol), 1-bromohexadecane(5.18 g, 17.0 mmol), K₂CO₃ (9.37 g, 67.8 mmol), CH₃CN (100 mL) yielded6.30 g (86.5%) of a white solid. ¹H NMR (CDCl3, 600 MHz) δ: 8.26 (t,3J=1.4 Hz, 1H, Ar—H), 7.74 (d, ₃J=1.5 Hz, 2H, Ar—H), 4.03 (t, ³J=6.5 Hz,2H, O—CH ₂), 3.94 (s, 6H, COO—CH ₃), 1.80 (p, ³J=7.3 Hz, 2H, OCH₂CH ₂),1.46 (p, ³J=7.6 Hz, 2H, OCH₂CH₂CH ₂), 1.26 (m, 24H), 0.88 (t, ³J=6.8,3H, CH₂—CH ₃.

1-Hexadecyloxy-3,5-bis(hydroxymethyl)benzene (E10)

The product was generated using general protocol E usingdimethyl-5-hexadecyloxyisophthalate (D10, 6.30 g), lithium aluminumhydride (1.27 g, 33.5 mmol), Et₂O (150 mL) yielded 2.06 g (37.9%) of awhite solid. ¹H NMR (CDCl₃, 400 MHz) δ: 6.93 (s, 1H, Ar—H), 6.85 (s, 2H,Ar—H), 4.67 (d, ³J=5.9 Hz, 4H, Ar—CH ₂—OH), 3.97 (t, 6.4 Hz, 2H, O—CH₂), 1.78 (p, ³J=7.6 Hz, 2H, OCH₂CH ₂), 1.66 (t, ³J=6.0 Hz, 2H, CH₂—OH),1.45 (p, ³J=7.6 Hz, 2H, O CH₂CH₂CH ₂), 1.26 (m, 24H), 0.88 (t, ³J=6.6,3H, CH₂—CH ₃).

1-hexadecyloxy-3,5-bis(bromomethyl)benzene (B10)

The product was generated via general protocol F using1-hexadecyloxy-3,5-bis(hydroxymethyl)benzene (E10), PBr₃, and THF. Aftercolumn chromatography (hexanes, then ramped to 2% EtOAc in hexanes), thereaction yielded 1.20 g (44%) of a white solid. ¹H NMR (CDCl₃, 400 MHz)δ: (s, 1H, Ar—H), 6.85 (s, 2H, Ar—H), 4.43 (s, 4H, Ar—CH ₂—Br), 3.96 (t,³J=6.4 Hz, 2H, O—CH ₂), 1.77 (p, ³J=7.4 Hz, 2H, OCH₂CH ₂), 1.45 (p,³J=7.5 Hz, 2H, O CH₂CH₂CH ₂), 1.26 (m, 24H), 0.88 (t, ³J=6.8, 3H, CH₂—CH₃).

1-hexadecyloxy-3,5-bis[(trimethlammonoum bromide)methyl]benzene (10)(3,5-C16)

The product was generated via general protocol C using1-hexadecyloxy-3,5-bis(bromomethyl)benzene (B10, 1.03 g, 2.04 mmol),NMe₃ (33% wt solution in EtOH, 1.95 mL, 8.17 mmol) and EtOH (150 mL).After recrystallization (EtOH/Et₂O) and drying the reaction produced0.723 g (77.2%) of a white solid, mp=208° C. (dec). ¹H NMR (D₂O, 400MHz) δ: (s, 1H, Ar—H), 7.33 (s, 2H, Ar—H), 4.61 (s, 4H, Ar—CH ₂—N), 4.10(t, 6.4 Hz, 2H, O—CH ₂), 3.18 (s, 6H, NH₃), 1.82 (p, ³J=7.4 Hz, 2H,OCH₂CH ₂), 1.50 (p, ³J=7.6 Hz, 2H, O CH₂CH₂CH ₂), 1.29 (m, 24H), 0.90(t, ³J=6.6, 3H, CH₂—CH ₃). ¹³C NMR (CD₃OD, 150 MHz) δ: 161.47, 161.52,130.21, 122.32, 69.91, 69.70, 53.51, 33.07, 30.79, 30.75, 30.74, 30.55,30.47, 30.33, 27.17, 23.73, 14.44. Anal. Calcd for C₃₀H₅₈OBr₂N₂: C,57.87; H, 9.39; N, 4.50. Found: C, 57.80; H, 9.58; N, 4.49.

A11-A12, B11-B12, 11-12 (MC10, MC14).

Synthetic details and analyses of these compounds have previously beenpublished. [Roszak, K. Z.; Torcivia, S. L.; Hamill, K. M.; Hill, A. R.;Radloff, K. R.; Crizer, D. M.; Middleton, A. M.; Caran, K. L. J. ColloidInterface Sci., 2009, 331, 560-5641.]

Synthesis of Pyridinium Compounds. Menschutkin Reaction:

The 1-hexadecloxy-3,5-bis(bromomethyl)benzene (1 equiv) was suspended inabsolute ethanol in a 500 mL 2-neck round bottom flask, which wasstirred with a magnetic stir bar under nitrogen. Pyridine (4 equiv, 99%,ACROS) was added with a syringe, and the reaction was refluxed in an oilbath overnight. The flask was then opened, and the excess pyridine andEtOH were allowed to evaporate under the continued flow of N₂. Theremaining solvent was removed under vacuum, and the crude product wasrecrystallized from hot EtOH and Et₂O.

3,5-C16 (pyridinium)

yielded 0.112 g (66.0%) ¹H NMR (D₂O, ³J=6.1 Hz, 400 MHz) d: 9.10 (d, 4H,Pyr-H), 8.63 (t, 2H, ³J=7.8 Hz Pyr-H), 8.14 (t, 4H, ³J=7.1 Hz Pyr-H),7.28 (s, 1H, Ar—H), 7.17 (s, 2H, Ar—H), 5.84 (s, 4H, Ar—CH ₂—N), 4.01(t, 6.4 Hz, 2H, O—CH ₂), 1.76 (p, ³J=7.3 Hz, 2H, OCH₂CH ₂), 1.45 (p,³J=7.8 Hz, 2H, O CH₂CH₂CH ₂), 1.29 (m, 24H), 0.90 (t, ³J=6.8, 3H, CH₂—CH₃).

Mesitylene-Pyridinium,Pyridinium,Pyridinium [M-P,P,P] (21)

1,3,5-Tribromomethylbenzene (0.1 g, 0.280 mmol) was dissolved in EtOH (5mL). Pyridine (0.091 mL, 1.12 mmol) was added and the reaction wasallowed to run at reflux overnight. Additional pyridine (0.046 mL, 0.56mmol) was added to and the reaction was allowed to run at refluxovernight once more. Upon concentration of the reaction mixture, theprecipitate was stirred in boiling acetone (˜10 mL) for five minutes andsubsequently filtered hot. After vacuum drying, the reaction produced0.100 g (60.5%) of 1.

Mesitylene-12,12,12[M-12,12,12] (22)

1,3,5-Tribromomethylbenzene (0.1 g, 0.280 mmol) was dissolved in EtOH (5mL). N, N-Dimethyldodecylamine (0.250 mL, 0.924 mmol) was added and thereaction was allowed to run at reflux overnight. Upon concentration ofthe reaction mixture, the precipitate was stirred in acetone (˜10 mL) atroom temperature for ten minutes. The precipitate was then filtered andvacuum dried.

Mesitylene-14,14,14 [M-14,14,14] (23)

1,3,5-Tribromomethylbenzene (0.1 g, 0.280 mmol) was dissolved in EtOH (5mL). N, N-Dimethyltetradecylamine (0.280 mL, 0.924 mmol) was added andthe reaction was allowed to run at reflux overnight. Upon concentrationof the reaction mixture, the precipitate was stirred in acetone (˜10 mL)at room temperature for ten minutes. The precipitate was then filteredand vacuum dried. The reaction produced 0.246 g (82.4%).

Mesitylene-16,16,16 [M-16,16,16] (24)

1,3,5-Tribromomethylbenzene (0.1 g, 0.280 mmol) was dissolved in EtOH (5mL). N, N-Dimethylhexadecylamine (0.311 mL, 0.924 mmol) was added andthe reaction was allowed to run at reflux overnight. Upon concentrationof the reaction mixture, the precipitate was stirred in acetone (˜10 mL)at room temperature for ten minutes. The precipitate was then filteredand vacuum dried. The reaction produced 0.235 g (87.7%).

Intermediate 1 (I1).

1,3,5-Tribromomethylbenzene (1.0 g, 2.8 mmol) was dissolved in acetone(50 mL). Pyridine (0.453 mL, 5.6 mmol) was added and the reaction wasallowed to run at room temperature overnight. The reaction precipitatewas filtered and subsequently stirred with acetone (˜100 mL) for tenminutes. After filtration of the insoluble product, the reactionproduced 0.9425 g (77.2%).

Mesitylene-Pyridinium,8,8 [M-P,8,8] (25)

I1 (0.1 g, 0.229 mmol) was dissolved in EtOH (5 mL). N,N-Dimethyloctylamine (0.104 mL, 0.505 mmol) was added and the reactionwas allowed to run at reflux for six hours. Upon concentration of thereaction mixture, the precipitate was stirred in acetone (˜10 mL) atroom temperature for ten minutes. The precipitate was then filtered anddried in vacuo.

Mesitylene-Pyridinium,10,10 [M-P,10,10] (26)

I1 (0.1 g, 0.229 mmol) was dissolved in EtOH (5 mL). N,N-Dimethyldecylamine (0.120 mL, 0.505 mmol) was added and the reactionwas allowed to run at reflux for six hours. Upon concentration of thereaction mixture, the precipitate was stirred in acetone (˜10 mL) atroom temperature for ten minutes. The precipitate was then filtered anddried in vacuo.

Mesitylene-Pyridinium,12,12 [M-P,12,12] (27)

I1 (0.25 g, 0.573 mmol) was dissolved in EtOH (5 mL). N,N-Dimethyldodecylamine (0.341 mL, 1.26 mmol) was added and the reactionwas allowed to run at reflux overnight. Upon concentration of thereaction mixture, the precipitate was stirred in acetone (˜10 mL) atroom temperature for ten minutes. The precipitate was then filtered anddried in vacuo. The reaction produced 0.383 g (77.5%).

Mesitylene-Pyridinium,14,14 [M-P,14,14] (28)

I1 (0.50 g, 1.15 mmol) was dissolved in EtOH (5 mL). N,N-Dimethyltetradecylamine (0.766 mL, 2.52 mmol) was added to thereaction mixture and refluxed overnight. Upon concentration of thereaction mixture, the precipitate was stirred in acetone (˜10 mL) atroom temperature for ten minutes. The precipitate was then filtered anddried in vacuo. The reaction produced 0.920 g (87.0%).

Mesitylene-Pyridinium,16,16 [M-P,16,16] (29)

I1 (0.25 g, 0.573 mmol) was dissolved in EtOH (5 mL). N,N-Dimethyldodecylamine (0.424 mL, 1.26 mmol) was added to the reactionmixture and refluxed overnight. Upon concentration of the reactionmixture, the precipitate was stirred in acetone (˜10 mL) at roomtemperature for ten minutes. The precipitate was then filtered and driedin vacuo. The reaction produced 0.512 g (91.6%).

ortho-Xylylene-14,14 [oX-14,14] (30)

o-Xylylene dibromide (0.5 g, 1.89 mmol, 1 equiv) was reacted underreflux with N,N-dimethyltetradecylamine (1.15 mL, 3.79 mmol, 2 equiv) inethanol (10 mL) overnight in 80° C. oil bath. The reaction was taken offthe heat and the precipitate was filtered. The crude product wasrecrystallized from H₂O, yielding a white product. ¹H NMR (DMSO-d6, 400MHz) δ: 7.73 (m, 4H, Ar—H), 4.74 (s, 4H, Ar—CH ₂), 3.39 (m, 4H, N—CH ₂),2.92 (s, 12H, N—CH ₃), 1.76 (br, 4H, N—CH₂—CH₂), 1.24 (br, 44H,N—CH₂—CH₂—(CH ₂)₁₁, 0.86 (t, 6H, N—(CH₂)₁₃—CH ₃).

meta-Xylylene-14,14 [mX-14,14] (31)

meta-Xylylene dibromide (0.6 g, 2.3 mmol, 1 equiv) andN,N-dimethyltetradecylamine (2.1 mL, 6.9 mmol, 3 equiv) were combined indry THF (50 mL). The reaction was run under reflux for 4 h. Uponcompletion, the reaction was taken off the heat and the crudeprecipitate was filtered. The product was recrystallized fromchloroform/acetone. The recrystallized white solid product was removedby filtration and then dried (1.132 g (65.9%). ¹H NMR (CDCl₃, 400 MHz)δ: 8.89 (s, 1H, Ar—H), 7.90 (d, 2H, Ar—H), 7.51 (t, 1H, Ar—H), 5.08 (s,4H, Ar—CH ₂), 3.57 (m, 4H, N—CH ₂), 3.26 (s, 12H, N—CH₃), 1.85 (m, 4H,N—CH₂—CH ₂), 1.26 (br, 44H, N—CH₂—CH₂—(CH₂)₁₁, 0.88 (t, 6H,N—(CH₂)₁₃—CH₃). ¹³C NMR (DMSO-d6, 150 MHz) δ: 138.49, 135.23, 129.83,128.53, 66.46, 64.94, 49.79, 31.80, 29.58, 29.56, 29.54, 29.53, 29.42,29.36, 29.24, 29.22, 26.32, 22.91, 22.57, 14.01.

para-Xylylene-14,14[pX-14,14] (32)

para-Xylylene dibromide (0.6 g, 2.3 mmol, 1 equiv) andN,N-dimethyltetradecylamine (2.1 mL, 6.9 mmol, 3 equiv) were combined indry THF (50 mL). The reaction was run under reflux for 4 h. Uponcompletion, the reaction was taken off the heat and the crudeprecipitate was filtered. The product was recrystallized fromchloroform/acetone yielding a white solid product.

ortho-Xylylene monopyridinium monobromide (33)

o-xylylene dibromide (15.00 g, 0.05682 mol, 3 equiv), pyridine (1.532mL, 0.01894 mol, 1 equiv) were dissolved in acetone (100 mL), and thereaction was allowed to run at reflux overnight. The reaction was takenoff the heat and the precipitate was removed by filtration andsubsequently recrystallized from acetonitrile yielding a whiteprecipitate (4.81 g (74.0%). ¹H NMR (DMSO, 400 MHz) δ: 9.18 (d, 2H,Pyr-H), 8.68 (t, 1H, Pyr-H), 8.22 (t, 2H, Pyr-H), 7.59 (s, 1H, Ar—H),7.45 (m, 2H, Ar—H), 7.33 (d, 1H, Ar—H), 6.17 (s, 2H, Ar—CH₂), 4.98 (s,2H, Ar—CH ₂). ¹³C NMR (DMSO, 100 MHz) δ: 146.18, 145.14, 137.20, 132.24,131.65, 130.47, 130.03, 129.66, 128.34, 59.95, 31.79.

meta-Xylylene monopyridinium monobromide (34)

m-xylylene dibromide (15.00 g, 0.05682 mol, 3 equiv), pyridine (1.532mL, 0.01894 mol, 1 equiv) were dissolved in acetone (50 mL), and thereaction was allowed to run at reflux overnight. The reaction was takenoff the heat and the precipitate was removed by filtration andsubsequently recrystallized from acetonitrile yielding a whiteprecipitate (5.15 g, 79.3%). ¹H NMR (DMSO, 400 MHz) δ: 9.24 (d, 2H,Pyr-H), 8.65 (t, 1H, Pyr-H), 8.21 (t, 2H, Pyr-H), 7.61 (s, 1H, Ar—H),7.47 (m, 3H, Ar—H), 5.90 (s, 2H, Ar—CH ₂), 4.69 (s, 2H, Ar—CH₂). ¹³C NMR(DMSO, 100 MHz) δ: 146.08, 144.87, 139.08, 134.77, 130.21, 129.60,129.47, 128.72, 128.51, 62.65, 33.68.

ortho-Xylylene-Pyridinium,14 [oX-P,14] (35)

13 (1.00 g, 2.92 mmol, 1 equiv) and N,N-dimethyltetradecylamine (1.062mL, 3.498 mmol, 1.2 equiv) were dissolved in ethanol (85 mL) and allowedto react under reflux overnight. The reaction was taken off the heat andthe precipitate was removed by filtration. The crude product wassuspended in acetone for about 20 min. The product was then removed byvacuum filtration, yielding a white solid (0.959 g, 56.3%). ¹H NMR(DMSO, 400 MHz) δ: 9.15 (d, 2H, Pyr-H), 8.70 (t, 1H, Pyr-H), 8.23 (t,2H, Pyr-H), 7.74 (m, 1H, Ar—H), 7.57 (m, 2H, Ar—H), 7.25 (m, 1H, Ar—H),6.31 (s, 2H, Ar—CH ₃), 4.92 (s, 2H, Ar—CH ₃), 3.49 (m, 2H, N—CH ₂), 3.03(s, 6H, N—CH₃), 1.81 (m, 3H, N—CH₂—CH ₂), 1.25 (br, 22H, N—CH₂—CH₂—(CH₂)₁₁), 0.85 (t, N—CH₂—CH₂—(CH₂)₁₁—CH ₃).

meta-Xylylene-Pyridinium,14 [mX-P,14] (36)

13 (0.622 g, 1.81 mmol, 1 equiv) and N,N-dimethyltetradecylamine (0.66mL, 2.2 mmol, 1.2 equiv) were dissolved in ethanol (85 mL) and allowedto react under reflux overnight. The reaction was taken off the heat andthe precipitate was removed by filtration. The crude product wasrecrystallized from acetone, yielding a white solid (0.728 g, 68.7%).

1-tetradecyloxy-2,5-bis(4,4′-bipyridinium)benzene, bis-bromide salt (41)1-tetradecyloxy-2,5-dimethylbenzene

1-bromotetradecane (44.4 mL, 0.149 mol), 2,5-dimethylphenol (20.0 g,0.164 mol) and K₂CO₃ (82.4 g, 0.596 mol) were combined in a round bottomflask in acetonitrile (200 mL). The flask was equipped with a stir-bar,and a water-cooled condenser and protected under a nitrogen atmosphere.The mixture was heated at reflux for overnight. Synthetic progress wasmonitored by the disappearance of the signal (triplet) from the C1hydrogens on the alkyl bromide (˜3.5 ppm) in ¹H NMR. Upon completion,the reaction was removed from heat and excess K₂CO₃ was removed bygravity filtration (rinsed with CH₂Cl₂). The crude product wasconcentrated in vacuo and the resulting material was dissolved inCH₂Cl₂, washed with 1 M NaOH (2×, to remove unreacted 2,5-dimethylphenolor p-cresol), dH₂O (2×) and brine (1×), dried over Na₂SO₄, gravityfiltered, and concentrated in vacuo to remove the solvent. The materialwas of sufficient purity (by ¹H and ¹³C NMR) to be used in thesubsequent reactions. Yielded 41.0 g (86%) of an off-white solid,mp=36.9-37.9° C. ¹H NMR (CDCl₃, 400 MHz) δ: 7.01 (d, ³J=7.4 Hz, 1H,Ar—H); 6.66 (d, ³J=7.6 Hz, 1H, Ar—H); 6.65 (s, 1H, Ar—H); 3.95 (t,³J=6.5 Hz, 2H, OCH ₂); 2.33 (s, 3H); 2.20 (s, 3H); 1.80 (m, 2H, OCH₂CH₂); 1.49 (m, 2H, OCH₂CH₂CH ₂); 1.42-1.26 (m, 20H); 0.90 (t, ³J=6.8 Hz,3H, CH₂CH ₃). ¹³C NMR (CDCl₃, 100 MHz) δ: 157.15, 136.40, 130.25,123.65, 120.53, 112.01, 67.88, 31.93, 29.70, 29.68, 29,66, 29.62, 29.42,29.40, 29.37, 26.16, 22.70, 21.40, 15.78, 14.11. HRMS (DART):[M+H]⁺=319.29460 (calcd for C₂₂H₃₉O 319.29954).

1-tetradecyloxy-2,5-bis(bromomethyl)benzene

All glassware was dried in an oven overnight and flushed with N₂ (g)prior to use. A solution of 1-tetradecyloxy-2,5-dimethylbenzene (A14,15.0 g, 47.1 mmol) in carbon tetrachloride (510 mL) was prepared in around bottom flask equipped with a water-cooled condenser, magnetic stirbar and protected with under a nitrogen atmosphere. N-bromosuccinimide(99%, 16.9 g, 94.2 mmol), benzoyl peroxide (0.234 g, 0.942 mmol) wereadded and the reaction was run at room temperature under irradiationfrom a 300 W/82V tungsten halogen lamp. The product was generated viageneral protocol B using, and CCl₄ (510 mL). The reaction was run in 6equal batches at room temperature under irradiation for 30-60 min perbatch. Reaction progress was monitored by the disappearance of theinsoluble NBS at the bottom of the flask and the appearance of the lowerdensity insoluble succinimide byproduct that floated to the surface ofthe reaction mixture. After filtration of the succinimide, the solventwas removed in vacuo. After two recrystallizations of the combined crudeproduct from n-hexane, 5.21 g (23%) of the pure product was obtained asa white solid, mp=64.0-64.2° C. ¹H NMR (CDCl₃, 600 MHz) δ: 7.28 (d,³J=7.6 Hz, 1H, Ar—H); 6.92 (dd, ³J=7.6 Hz, ⁴J=1.6 Hz, 1H, Ar—H); 6.88(d, ⁴J=1.6 Hz, 1H, Ar—H); 4.54 (s, 2H); 4.45 (s, 2H); 4.04 (t, ³J=6.4Hz, 2H, OCH ₂); 1.84 (m, 2H, OCH₂CH ₂); 1.51 (m, 2H, OCH₂CH₂CH ₂);1.40-1.21 (m, 20H); 0.88 (t, ³J=7.0 Hz, 3H, CH₂CH ₃). ¹³C NMR (CDCl₃ 150MHz) δ: 157.11, 139.73, 131.02, 126.50, 120.92, 112.27, 68.31, 33.36,31.92, 29.68, 29.67, 29.65, 29.59, 29.58, 29.36, 29.33, 29.15, 28.48,26.06, 22.69, 14.13. HRMS (DART): [M−Br]³⁰=395.19344 (calcd forC₂₂H₃₆BrO 395.19440).

1-tetradecyloxy-2,5-bis(4,4′-bipyridinium)benzene, bis-bromide salt

4,4′bipyridine (98.9 mg, 0.634 mmol) was dissolved in CH₃CN (10 mL) andbrought to reflux in a round bottom flask equipped with anitrogen-protected, water-cooled reflux condenser. A solution of1-tetradecyloxy-2,5-bis(bromomethyl)benzene (100 mg, 0.210 mmol)dissolved in CH₃CN (2.5 mL) and CHCl₃ (5.0 mL) was added dropwise to thesolution of 4,4′-bipyridine. The reflux condenser was switched out for adistillation head and condenser ˜1 h in order to distill off the CHCl₃.After this, the reflux condenser was switched back in and the reactionwas allowed to run at reflux for 5 h. Upon cooling, the solid productprecipitated from solution. It was collected by filtration and washedwith cold CH₃CN, and subsequently recrystallized from CH₃CN and dried,producing a yellow crystalline product (128 mg, 0.162 mmol, 77%). ¹NMR(CD₃OD, 400 MHz) δ: 9.22 (d, ³J=6.4 Hz, 2H, Ar—H); 9.10 (d, ³J=6.4 Hz,2H, Ar—H); 8.82 (m, 4H, Ar—H), 8.53 (m, 4H, Ar—H); 7.97 (m, 4H, Ar—H);7.74 (d, ³J=7.4 Hz, 1H, Ar—H); 7.38 (s, 1H, Ar—H); 7.28 (d, 7.4 Hz, 1H,Ar—H); 5.93 (s, 2H, Ar—CH ₂—N); 5.89 (s, 2H, Ar—CH ₂—N); 4.09 (t, 2H,³J=6.4 Hz, OCH ₂); 1.73 (m, 2H, OCH₂CH ₂); 1.35-1.13 (m, 22H), 0.89 (t,6.6 Hz, CH₂CH ₃).

What is claimed is:
 1. A pharmaceutical composition comprising acompound of formula I:

wherein R₁, R₂, and R₃, and/or R₅, R₆, and R₇, and/or R₈, R₉, and R₁₀,together with the nitrogen atom to which they are attached, form apyridinium or pyridyl-substituted pyridinium; m is 1; n is 1; p is 0; tis 1; R₄ is C₈₋₂₂alkyl; and X is halogen or tartrate; and apharmaceutically acceptable carrier or diluent.
 2. The pharmaceuticalcomposition of claim 1, wherein the compound of formula I is one of


3. The pharmaceutical composition of claim 1, wherein the compound offormula I is


4. The pharmaceutical composition of claim 1, wherein R₄ is—C₁₀₋₁₈alkyl.
 5. The pharmaceutical composition of claim 4, wherein R₄is —C₁₀H₂₁.
 6. The pharmaceutical composition of claim 4, wherein R₄ is—C₁₂H₂₅.
 7. The pharmaceutical composition of claim 4, wherein R₄ is—C₁₄H₂₉.
 8. The pharmaceutical composition of claim 4, wherein R₄ is—C₁₆H₃₃.
 9. The pharmaceutical composition of claim 4, wherein R₄ is—C₁₈H₃₇.
 10. The pharmaceutical composition of claim 1, wherein thecompound of formula I is selected from the following Table:

Comp. No. Substitution R₄ 15 2, 3 —C₁₄H₂₉ 40 3, 5 —C₁₄H₂₉ 16 3, 5 —C₁₆H₃₃.


11. The pharmaceutical composition of claim 1, wherein the compound offormula I is


12. The pharmaceutical composition of claim 1, wherein X is Br.